The objective of this proposal is to characterize hexose transport in Chinese hamster ovary cells in culture utilizing techniques of biochemistry and genetics. Employing a rapid assay, uptake of the following hexoses will be measured; 3-0-metabolizable sugar; D-glucose in ATP-depleted cells; and D-galactose in galactokinase deficient cells. Uptake under these non-metabolizing conditions can clearly be ascribed to transport of hexoses rather than to their metabolism. If initial velocities can not be accurately measured when the uptake is less than 10% of the equilibrium value, data from following the reaction until equilibrium will be used to calculate kinetic parameters from an integrated rate equation. The activation of hexose transport after glucose deprivation and the role of sulfhydryl groups in this activation will be investigated. Three independent methods for isolating mutants in hexose transport have been described: resistance to killing by glucose analogues; resistance to killing by 3H-hexoses; and resistance to killing by inhibitors of glucose transport. Characterization of these mutants will distinguish between a structural gene mutation and a regulatory gene mutation. Furthermore, mutant characterization will provide new information about the hexose carrier in terms of its regulation, kinetic parameters, number of carriers, substrate specificity, and the identification of membrane proteins involved in hexose transport.